IGEM:UNAM-Genomics Mexico/2009/Notebook/Collaborations/2010/08/25

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Shipment to Edinburgh team

Hello Mexico!

Thank you very much for the papers and your last e-mail with all the
links- it's really useful.

I've recently tried to fix our LovTap by introducing missing base,
subculturing, minipreping etc. I've submitted the newst clones to
sequencing and hopefully tomorrow I'll get the results back, so we can
retransform the cells and get to the characterization assay.

I didn't have a chance to work on the pcb promoter, I've passed your
e-mail to Will and hopefully he will get back to you soon about this.

Also, as I'm going for a holidays on Friday, Maria will take over the
LovTap and other experimetns from me. Will is mainly working with
Luciferases. I'll forward emails to them if you send them to me,if I
can't answer myself. But feel free to still contact me.

Once we have our parts ready we'll send them to you- the team will
keep you updated. Thank you for your parts.

Good luck with all your work. Again- if you have any questions I'm happy to answer or you can contact
Maria or Will- here are their e-mail addresses:

Maria- M.T.Kowal@sms.ed.ac.uk

Will - W.G.P.M.Rostain@sms.ed.ac.uk

Good luck with the lab work!

Marta

Answer quoting chernand@lcg.unam.mx

Hi Edinburgh Team!

We have been working during this week in order to have the shipment ready.
As the process of restriction enzyme assays, DNA gel extraction and
ligations of the synthesized constructions is very slow, taking into
account the experience working with LovTAP. We considered that the best
option will be to send you the PCR products from the parts synthesized.

We are planning to send you:

-CcaS&CcaR PCR product.
-LuxY PCR product.
-LovTAP PCR product.

All these sequences have the prefix and suffix sequences from the biobrick
assembly protocol in the ends.

If LovTAP ligation with plasmid pSB1C3 is successful, we are going to send
you it, too.

The experimental details of the PCR reactions and LovTAP gel extraction
and ligation can be found in the next links:

Explore the logbook for more information and check it continuously for
future updates.

LuxAB

Augusto had to repeat the LuxAB extraction from Vibrio fischeri genomic
DNA, the primers that he is using already include the prefix and suffix
sequences to assemble the LuxAB amplified product with the biobrick
standard protocol. We could send you this PCR product if you were
interested.

Luciferase

One strain harboring the luciferase mutant S284T, was analyzed in a
preliminary test using luciferin and waiting 30 seconds of exposition, but
there was no bioluminescence emission. The same happened with the WT
luciferase. I don’t have all the details of the experiment, but those were
the results. Mariana and Enrique will repeat the assay with other colonies
and inside dark room. If we have the functional mutant, we will send it
to you.

Blue promoter

The blue promoter inside plasmid pSB1C3 is ready, maybe we could send you
the promoter ligated to GFP, Jorge is checking the ligation.

We are planning to send the parts on Thursday or Friday.

During the weekend we are going to update the information of the
constructions that you will receive, thus you will get totally
familiarized with them. We remain at your disposition for any question.

For us, it will be great to receive:

-LovTAP Read out system 1

-Cph8 PCR

-Mutated Luciferase 356K

-The plasmid harboring the genes to produce PCB in E.coli. (If you get it
first)

If you have any suggestion or comment about the shipment, let us know. We
are delighted to help you.